The present invention relates to an electrophoresis apparatus for separatingly detecting DNA, RNA or the like, and more particularly to a wavelength dispersion electrophoresis apparatus which is well suited for detecting fluorescences from a sample labeled with a plurality of fluorophores having unequal wavelengths.
Heretofore, the determination of a base sequence on DNA has been performed in such a way that a separation pattern is transferred on a photograph by the use of autoradiography. However, there has been the difficulty that labor and time are expended besides the troublesome handling of radioactive elements. Therefore, note is taken of a system wherein a DNA fragment during electrophoresis separation is detected in real time through the fluorescence labeling of the DNA (L. M. Smith et al; Nature, Vol. 321, pp. 674-679 (1986)). With this method, while the specified end of the DNA is maintained, the fluorescence labeling is done at the other end or halfway so as to prepare four kinds of DNA fragment groups in each of which a base possessed by a nucleic acid at the other end is adenine (A), cytosine (C), thymine (T) or guanine (G). At this time, fluorophores of unequal emission wavelengths are used for labeling the respective base species. The four fragment groups are put together, and are subjected to the electrophoresis gel separation. The electrophoretic migration speed is higher for a shorter DNA fragment. Therefore, when a place at a fixed distance from a sample inlet is irradiated by a laser, the fragments pass through the irradiated region and give forth fluorescences successively from the shortest fragment. Since the emission wavelengths differ depending upon the base species, the base species are determined from the wavelengths. The lengths of the fragments can be determined from the migration times.
In order to discriminate the four kinds of fluorescences of unequal wavelengths, L. M. Smith et al rotate four sorts of filters of unequal transmission wavelengths by way of example. Besides, an apparatus which employs a flat plate type gel in order to permit the measurements of a plurality of samples is sold by Applied Biosystems Inc. in U.S. In this apparatus, a laser beam is fined to irradiate one point of a gel plane. Fluorescence emitted from here passes through a rotary filter, and is detected by a photosensitive portion. In measuring a large number of samples, information items are obtained in such a way that the gel plane is scanned on one straight line by interlocking laser irradiation positions and the detecting portion.
In the prior art employing the rotary filter, it has not been considered to increase the amount of the fluorescence which is received by the detecting portion. More specifically, when it is intended to detect the region of the surface of an electrophoretic plate in a range of 10 cm in a horizontal direction (a direction orthogonal to the direction of the electrophoresis), the period of time for which the laser irradiates each spot becomes 1/200 of one scanning time (usually, 1 second or so), assuming the width of the laser beam to be 0.5 mm. Moreover, since four filters rotate in correspondence with the four base species, the measurement period of time for one base species noticed is further quartered. Eventually, only about 1/800 of the whole measurement period of time is used for the measurement of one base species at one spot, so that a high sensitivity is not attained due to a small amount of fluorescence received.